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212 Scalp hypothermia as a preventive measure for chemotherapy-induced alopecia: A systematic review of the literature
Hair and Other Adnexal Structures | ABSTRACTS 210
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Developing an in vitro model to dissect reciprocal mesenchymal-epithelial signalling in the hair follicle C Platt and R Paus Inflammation and Repair, University of Manchester, Manchester, United Kingdom Current in vitro models of hair growth that use isolated human hair follicles, although highly physiologically relevant, require a significant amount of high quality, post-operative tissue. In contrast, the use of dissociated cells to model hair growth may lack the sophistication required to recreate the complex signalling networks present in the growing hair follicle. In this study we aimed to develop a heterotypic in vitro model to explore the reciprocal mesenchymal-epithelial signalling that regulates human hair growth and pigmentation. Our in vitro model consisted of plucked human scalp hair (epithelial component) co-cultured with human dermal papilla spheroids (DPS; mesenchymal component). Plucked scalp hair was obtained from healthy volunteers and dermal papilla cells were isolated from human scalp hair follicles obtained from transplant surgery. Our results showed that freshly plucked scalp follicles contained cells positive for Ki-67, GP100, tyrosinase activity and Masson Fontana. Positive staining was localised to the epithelial matrix. DPS cultured in hanging drops were viable after 48h in culture and expressed the inductive markers alkaline phosphatase and versican. Surprisingly, co-culture of plucked scalp hair with DPS for 48h, on a low adhesive substrate, resulted in attachment of DPS to the plucked scalp hair. Subsequent histological analysis revealed that DPS had surrounded and engulfed the epithelial matrix of the plucked scalp hair. Data from this study indicates that the epithelial and mesenchymal components of our model are phenotypically consistent with intact human scalp hair follicles, with regard to markers of cell proliferation, pigmentation and induction. In addition, the physical interaction of DPS and plucked hair, in co-culture, provides the basis for an in vitro model to study reciprocal signalling between hair follicle epithelium and mesenchyme. Future studies will use this model to explore whether the attached DPS stimulates epithelial cell proliferation and pigmentation in the co-cultured plucked scalp hair.
LRIG1 a regulator of ERBB signaling in skin during development and homeostasis C Hoesl1, M Schneider1, E Wolf1, R Wolf2 and M Dahlhoff1 1 Institute of Molecular Animal Breeding and Biotechnology, LMU Munich, Munich, Germany and 2 Klinik und Poliklinik fu¨r Dermatologie und Allergologie, LMU Mu¨nchen, Munich, Germany The leucine-rich repeats and immunoglobulin-like domains (LRIG) family includes transmembrane proteins known to be essential regulators of growth factor receptors like the epidermal growth factor receptor (EGFR/ERBB) family. As ERBBs are involved in cell proliferation, differentiation, death, motility, and adhesion, they are very versatile players during processes such as development, tissue homeostasis and tumorigenesis. The LRIG proteins are thought to regulate ERBBs and also other receptor tyrosine kinases. However, the role of LRIG proteins for regulatory feedback loops is not understood yet and requires further studies. To study LRIG1 in more detail, we generated doxycycline-inducible, skin-targeted (keratin 5 promoter-directed) transgenic (tg) mouse lines overexpressing LRIG1 using the TET-OFF system. As it is known that LRIG1-knockout mice develop a hyperplasia of the epidermis and psoriasis due to a hyperactive EGFR receptor, we anticipated a thinner epidermis in LRIG1 tg mice. Surprisingly, we could not obtain LRIG1 overexpressing mice in the expected rates. Only 9% (6/65) of all born mice were both LRIG1 and tTA positive and 83% (5/6) of these mice died immediately after birth. Only one double transgenic mouse survived for 13 days. The hair coat development of this animal was delayed and it showed reduced hair growth, hyperkeratosis, utricle development and a thicker epidermis compared with control siblings. Initial investigations in LRIG1 tg mice at day P0 suggest that the loricrin positive epidermal layer was increased in this animals. To study the phenotype of LRIG1 tg also in older animals, we suppressed the LRIG1 overexpression until birth by doxycycline treatment of pregnant females. Induction of the LRIG1 expression after birth by discontinued doxycycline application enables the survival of LRIG1 transgenic mice. These mice showed alopecia from three months of age with a significant thicker epidermis and utricles.
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Scalp hypothermia as a preventive measure for chemotherapy-induced alopecia: A systematic review of the literature V Shah1, T Wikramanayake1, GM DelCanto2, AV Granda1, A Lens1 and JJ Jimenez1 1 Department of Dermatology and Cutaneous Surgery, University of Miami Miller School of Medicine, Miami, FL and 2 Biochemistry and Molecular Biology, University of Miami, Miami, FL Chemotherapy-induced alopecia (CIA) is a psychologically devastating condition with serious impact on the well-being of patients undergoing treatment. The most prevalent preventive measure for CIA involves cooling the scalp before and during infusion of chemotherapeutic drugs, but despite its widespread use, there remains some controversy among practitioners over the efficacy of scalp hypothermia. For this reason, we sought to evaluate the efficacy of scalp cooling for the prevention of CIA. We performed a systematic search of the electronic databases PubMed, Google Scholar, the Cochrane Central Register of Controlled Trials (CENTRAL), and TRIP to identify original articles that evaluated scalp cooling for the treatment of CIA. Data were extracted from each study and consolidated into standardized tables. A total of 489 related citations were returned after the initial search. Following evaluation based upon inclusion/exclusion criteria, 7 randomized and 11 controlled trials were included in this systematic study, with 802 total patients receiving scalp-hypothermia therapy. Six of the 7 randomized clinical trials and all but one controlled clinical trial demonstrated significantly improved hair preservation following scalp cooling (p<0.01). Numerous factors have been shown to influence the effectiveness of scalp cooling, such as cooling time, temperature, and chemotherapeutic regimen. Overall, the results suggest that scalp hypothermia is a useful procedure for the treatment of CIA. However, the data show varying levels of success, and head-to-head comparisons are difficult, owing to significant technical variations between studies—e.g., in duration, chemotherapy regimens, and scalp-cooling techniques/devices. Further optimization of the factors that influence scalp cooling is imperative in order to implement more standardized treatment for maximum efficacy.
Whiter teeth without autophagy e deletion of Atg7 in K14-positive cells impairs the iron metabolism of the murine enamel epithelium S Sukseree1, H Rossiter1, UY Schwarze2, R Gruber2, E Tschachler1 and L Eckhart1 1 Department of Dermatology, Medical University of Vienna, Vienna, Austria and 2 Department of Oral Biology, Medical University of Vienna, Vienna, Austria Autophagy is an intracellular vesicular transport and lysosomal fusion mechanism that is active in many cell types upon exposure to stress or induction of differentiation. We have generated Atg7f/f K14-Cre mice in which the essential autophagy gene Atg7 is deleted in keratin K14-expressing cells. Here, we used this model, in comparison to wildtype mice, to investigate the role of Atg7 in ameloblasts, the cells that produce the enamel of teeth. The epithelium-specific deletion of Atg7 was compatible with the survival and enamel-forming ability of ameloblasts. However, the incisors, which grow throughout life, often suffered mechanical damage and developed secondary malformations in aging Atg7f/f K14-Cre mice. Histological studies, including Perls Prussian blue staining, of the maxillary jaw and incisors showed that the transport of iron from ameloblasts into the forming enamel was blocked in the absence of Atg7. While iron was deposited on the surface of normal enamel, it was aberrantly retained in enamel epithelial cells and subsequently taken up by macrophages in Atg7f/f K14-Cre mice. Due to the lack of iron in the enamel, the color of the incisors was white instead of the normal yellow. In conclusion, these results suggest that the autophagy regulator Atg7 plays an essential role in the normal function of murine ameloblasts. Moreover, this study demonstrates that transgenic mouse models for studies of the epidermis can have unexpected “off-target” phenotypes in other epithelia.
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Growth hormone-induced signaling: A novel, intrafollicular neuroendocrine control of human hair growth M Alam1, D Below1, R Clayton3, M Bertolini1, J Che´ret2 and R Paus3 1 Dermatology, University of Muenster, Muenster, Germany, 2 Dermatology, Monasterium Laboratory, Muenster, Germany and 3 Dermatology, University of Manchester, Manchester, United Kingdom Growth hormone (GH) and activation of its receptor (GHR) promote cell growth, proliferation, and differentiation either directly or via IGF-1 induction. While clinical case reports suggest that human hair follicles (HFs) may be GH-responsive, it is unknown whether GH exerts any functions in human HF biology. To explore this, we charted the expression of GH, GHR and growth hormone releasing hormone (GHRH), the hypothalamic neurohormone that stimulates pituitary GH expression and release, in human scalp HFs. So far, we have detected GH, GHR and GHRH protein expression in the ORS of human anagen VI HFs as well as in the sebaceous gland epithelium. qRT-PCR analysis revealed that human scalp HFs also transcribe GH and GHR mRNA, supporting that human HFs are an extrapituitary source of GH production. qRT-PCR analysis showed that GHR transcription decreases during HF regression (catagen HFs), whilst the level of the GH-inhibiting hormone, somatostatin (SST), was increased. Recombinant hGH (rGH, 100-200 ng/ml) induced premature catagen development ex vivo in female microdissected HFs, along with a significant increase of intrafollicular IGF-1 and TGFb2 protein immunoreactivity. qRT-PCR analysis showed that hGH (300 ng/ml) treated HFs, reduced transcript levels of GHRH, GHR and SST, while GH and IGF-1 mRNA levels were relatively unchanged. This suggests that the catagen-promoting effects of GH on human HFs are rather IGF-1 independent. Interestingly, scalp HFs stimulation with GHRH (300ng/ml) ex vivo up-regulated GH transcription, supporting that GH is indeed transcribed intrafollicularly (e.g. via GHRH). While we are still investigating how GHR silencing in human HFs ex vivo impacts on hair biology, our study already shows that GHR-mediated signaling by intrafollicularly generated GH is a major novel neuroendocrine regulator of human hair growth.
TLR2 and TLR4 activated SZ95 sebocytes promote inflammation prior to changing lipid metabolism at the level of gene expression D Torocsik1, S Poliska2, D Kovacs3, M Lova´szi3, Z Kovacs3, C Zouboulis4 and M Sta˚hle1 1 Unit of Dermatology and Venereology, Karolinska lnstitutet, Stockholm, Sweden, 2 Department of Biochemistry and Molecular Biology, Debrecen, Hungary, 3 Department of Dermatology, University of Debrecen, Debrecen, Hungary and 4 Departments of Dermatology, Venereology, Allergology and Immunology, Dessau Medical Center, Dessau, Germany Toll Like Receptor (TLR) 2 and 4 are central in the development of acne, however very little is known about their effect on gene expression regulation in sebocytes. In this work we therefore performed global gene expression profile analysis with functional clustering of the differentially regulated genes of TLR 2 (PAM3CSK4) and 4 (LPS) activated SZ95 sebocytes at early (6 hours) and late (24 hours) time points after treatment. Our results showed that both the TLR 2 and 4 activated sebocytes promoted inflammation in a similar manner already at the early time-point, with the regulation of genes involved in chemotaxis, wounding, cell proliferation, and a more complex cytokine and chemokine production than what was previously known. Moreover our data also identified a set of novel markers that has not been described yet for inflamed sebocytes such as SAA1 and 2, SERPINB 3 and 4, CCL20 and LCN2. Importantly lipid metabolism, the primary feature of sebocytes, was changed only at the later time point suggesting that sebocytes prioritize to exert an inflammatory phenotype in the presence of a danger signal. Our results underpin the complex role of sebocytes in inflammation and disease specific settings such as acne in which they could link lipid metabolism with inflammation via a prompt response at the level of gene expression regulation.
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Developing an in vitro model to dissect reciprocal mesenchymal-epithelial signalling in the hair follicle C Platt and R Paus Inflammation and Repair, University of Manchester, Manchester, United Kingdom Current in vitro models of hair growth that use isolated human hair follicles, although highly physiologically relevant, require a significant amount of high quality, post-operative tissue. In contrast, the use of dissociated cells to model hair growth may lack the sophistication required to recreate the complex signalling networks present in the growing hair follicle. In this study we aimed to develop a heterotypic in vitro model to explore the reciprocal mesenchymal-epithelial signalling that regulates human hair growth and pigmentation. Our in vitro model consisted of plucked human scalp hair (epithelial component) co-cultured with human dermal papilla spheroids (DPS; mesenchymal component). Plucked scalp hair was obtained from healthy volunteers and dermal papilla cells were isolated from human scalp hair follicles obtained from transplant surgery. Our results showed that freshly plucked scalp follicles contained cells positive for Ki-67, GP100, tyrosinase activity and Masson Fontana. Positive staining was localised to the epithelial matrix. DPS cultured in hanging drops were viable after 48h in culture and expressed the inductive markers alkaline phosphatase and versican. Surprisingly, co-culture of plucked scalp hair with DPS for 48h, on a low adhesive substrate, resulted in attachment of DPS to the plucked scalp hair. Subsequent histological analysis revealed that DPS had surrounded and engulfed the epithelial matrix of the plucked scalp hair. Data from this study indicates that the epithelial and mesenchymal components of our model are phenotypically consistent with intact human scalp hair follicles, with regard to markers of cell proliferation, pigmentation and induction. In addition, the physical interaction of DPS and plucked hair, in co-culture, provides the basis for an in vitro model to study reciprocal signalling between hair follicle epithelium and mesenchyme. Future studies will use this model to explore whether the attached DPS stimulates epithelial cell proliferation and pigmentation in the co-cultured plucked scalp hair.
LRIG1 a regulator of ERBB signaling in skin during development and homeostasis C Hoesl1, M Schneider1, E Wolf1, R Wolf2 and M Dahlhoff1 1 Institute of Molecular Animal Breeding and Biotechnology, LMU Munich, Munich, Germany and 2 Klinik und Poliklinik fu¨r Dermatologie und Allergologie, LMU Mu¨nchen, Munich, Germany The leucine-rich repeats and immunoglobulin-like domains (LRIG) family includes transmembrane proteins known to be essential regulators of growth factor receptors like the epidermal growth factor receptor (EGFR/ERBB) family. As ERBBs are involved in cell proliferation, differentiation, death, motility, and adhesion, they are very versatile players during processes such as development, tissue homeostasis and tumorigenesis. The LRIG proteins are thought to regulate ERBBs and also other receptor tyrosine kinases. However, the role of LRIG proteins for regulatory feedback loops is not understood yet and requires further studies. To study LRIG1 in more detail, we generated doxycycline-inducible, skin-targeted (keratin 5 promoter-directed) transgenic (tg) mouse lines overexpressing LRIG1 using the TET-OFF system. As it is known that LRIG1-knockout mice develop a hyperplasia of the epidermis and psoriasis due to a hyperactive EGFR receptor, we anticipated a thinner epidermis in LRIG1 tg mice. Surprisingly, we could not obtain LRIG1 overexpressing mice in the expected rates. Only 9% (6/65) of all born mice were both LRIG1 and tTA positive and 83% (5/6) of these mice died immediately after birth. Only one double transgenic mouse survived for 13 days. The hair coat development of this animal was delayed and it showed reduced hair growth, hyperkeratosis, utricle development and a thicker epidermis compared with control siblings. Initial investigations in LRIG1 tg mice at day P0 suggest that the loricrin positive epidermal layer was increased in this animals. To study the phenotype of LRIG1 tg also in older animals, we suppressed the LRIG1 overexpression until birth by doxycycline treatment of pregnant females. Induction of the LRIG1 expression after birth by discontinued doxycycline application enables the survival of LRIG1 transgenic mice. These mice showed alopecia from three months of age with a significant thicker epidermis and utricles.
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Scalp hypothermia as a preventive measure for chemotherapy-induced alopecia: A systematic review of the literature V Shah1, T Wikramanayake1, GM DelCanto2, AV Granda1, A Lens1 and JJ Jimenez1 1 Department of Dermatology and Cutaneous Surgery, University of Miami Miller School of Medicine, Miami, FL and 2 Biochemistry and Molecular Biology, University of Miami, Miami, FL Chemotherapy-induced alopecia (CIA) is a psychologically devastating condition with serious impact on the well-being of patients undergoing treatment. The most prevalent preventive measure for CIA involves cooling the scalp before and during infusion of chemotherapeutic drugs, but despite its widespread use, there remains some controversy among practitioners over the efficacy of scalp hypothermia. For this reason, we sought to evaluate the efficacy of scalp cooling for the prevention of CIA. We performed a systematic search of the electronic databases PubMed, Google Scholar, the Cochrane Central Register of Controlled Trials (CENTRAL), and TRIP to identify original articles that evaluated scalp cooling for the treatment of CIA. Data were extracted from each study and consolidated into standardized tables. A total of 489 related citations were returned after the initial search. Following evaluation based upon inclusion/exclusion criteria, 7 randomized and 11 controlled trials were included in this systematic study, with 802 total patients receiving scalp-hypothermia therapy. Six of the 7 randomized clinical trials and all but one controlled clinical trial demonstrated significantly improved hair preservation following scalp cooling (p<0.01). Numerous factors have been shown to influence the effectiveness of scalp cooling, such as cooling time, temperature, and chemotherapeutic regimen. Overall, the results suggest that scalp hypothermia is a useful procedure for the treatment of CIA. However, the data show varying levels of success, and head-to-head comparisons are difficult, owing to significant technical variations between studies—e.g., in duration, chemotherapy regimens, and scalp-cooling techniques/devices. Further optimization of the factors that influence scalp cooling is imperative in order to implement more standardized treatment for maximum efficacy.
Whiter teeth without autophagy e deletion of Atg7 in K14-positive cells impairs the iron metabolism of the murine enamel epithelium S Sukseree1, H Rossiter1, UY Schwarze2, R Gruber2, E Tschachler1 and L Eckhart1 1 Department of Dermatology, Medical University of Vienna, Vienna, Austria and 2 Department of Oral Biology, Medical University of Vienna, Vienna, Austria Autophagy is an intracellular vesicular transport and lysosomal fusion mechanism that is active in many cell types upon exposure to stress or induction of differentiation. We have generated Atg7f/f K14-Cre mice in which the essential autophagy gene Atg7 is deleted in keratin K14-expressing cells. Here, we used this model, in comparison to wildtype mice, to investigate the role of Atg7 in ameloblasts, the cells that produce the enamel of teeth. The epithelium-specific deletion of Atg7 was compatible with the survival and enamel-forming ability of ameloblasts. However, the incisors, which grow throughout life, often suffered mechanical damage and developed secondary malformations in aging Atg7f/f K14-Cre mice. Histological studies, including Perls Prussian blue staining, of the maxillary jaw and incisors showed that the transport of iron from ameloblasts into the forming enamel was blocked in the absence of Atg7. While iron was deposited on the surface of normal enamel, it was aberrantly retained in enamel epithelial cells and subsequently taken up by macrophages in Atg7f/f K14-Cre mice. Due to the lack of iron in the enamel, the color of the incisors was white instead of the normal yellow. In conclusion, these results suggest that the autophagy regulator Atg7 plays an essential role in the normal function of murine ameloblasts. Moreover, this study demonstrates that transgenic mouse models for studies of the epidermis can have unexpected “off-target” phenotypes in other epithelia.
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Growth hormone-induced signaling: A novel, intrafollicular neuroendocrine control of human hair growth M Alam1, D Below1, R Clayton3, M Bertolini1, J Che´ret2 and R Paus3 1 Dermatology, University of Muenster, Muenster, Germany, 2 Dermatology, Monasterium Laboratory, Muenster, Germany and 3 Dermatology, University of Manchester, Manchester, United Kingdom Growth hormone (GH) and activation of its receptor (GHR) promote cell growth, proliferation, and differentiation either directly or via IGF-1 induction. While clinical case reports suggest that human hair follicles (HFs) may be GH-responsive, it is unknown whether GH exerts any functions in human HF biology. To explore this, we charted the expression of GH, GHR and growth hormone releasing hormone (GHRH), the hypothalamic neurohormone that stimulates pituitary GH expression and release, in human scalp HFs. So far, we have detected GH, GHR and GHRH protein expression in the ORS of human anagen VI HFs as well as in the sebaceous gland epithelium. qRT-PCR analysis revealed that human scalp HFs also transcribe GH and GHR mRNA, supporting that human HFs are an extrapituitary source of GH production. qRT-PCR analysis showed that GHR transcription decreases during HF regression (catagen HFs), whilst the level of the GH-inhibiting hormone, somatostatin (SST), was increased. Recombinant hGH (rGH, 100-200 ng/ml) induced premature catagen development ex vivo in female microdissected HFs, along with a significant increase of intrafollicular IGF-1 and TGFb2 protein immunoreactivity. qRT-PCR analysis showed that hGH (300 ng/ml) treated HFs, reduced transcript levels of GHRH, GHR and SST, while GH and IGF-1 mRNA levels were relatively unchanged. This suggests that the catagen-promoting effects of GH on human HFs are rather IGF-1 independent. Interestingly, scalp HFs stimulation with GHRH (300ng/ml) ex vivo up-regulated GH transcription, supporting that GH is indeed transcribed intrafollicularly (e.g. via GHRH). While we are still investigating how GHR silencing in human HFs ex vivo impacts on hair biology, our study already shows that GHR-mediated signaling by intrafollicularly generated GH is a major novel neuroendocrine regulator of human hair growth.
TLR2 and TLR4 activated SZ95 sebocytes promote inflammation prior to changing lipid metabolism at the level of gene expression D Torocsik1, S Poliska2, D Kovacs3, M Lova´szi3, Z Kovacs3, C Zouboulis4 and M Sta˚hle1 1 Unit of Dermatology and Venereology, Karolinska lnstitutet, Stockholm, Sweden, 2 Department of Biochemistry and Molecular Biology, Debrecen, Hungary, 3 Department of Dermatology, University of Debrecen, Debrecen, Hungary and 4 Departments of Dermatology, Venereology, Allergology and Immunology, Dessau Medical Center, Dessau, Germany Toll Like Receptor (TLR) 2 and 4 are central in the development of acne, however very little is known about their effect on gene expression regulation in sebocytes. In this work we therefore performed global gene expression profile analysis with functional clustering of the differentially regulated genes of TLR 2 (PAM3CSK4) and 4 (LPS) activated SZ95 sebocytes at early (6 hours) and late (24 hours) time points after treatment. Our results showed that both the TLR 2 and 4 activated sebocytes promoted inflammation in a similar manner already at the early time-point, with the regulation of genes involved in chemotaxis, wounding, cell proliferation, and a more complex cytokine and chemokine production than what was previously known. Moreover our data also identified a set of novel markers that has not been described yet for inflamed sebocytes such as SAA1 and 2, SERPINB 3 and 4, CCL20 and LCN2. Importantly lipid metabolism, the primary feature of sebocytes, was changed only at the later time point suggesting that sebocytes prioritize to exert an inflammatory phenotype in the presence of a danger signal. Our results underpin the complex role of sebocytes in inflammation and disease specific settings such as acne in which they could link lipid metabolism with inflammation via a prompt response at the level of gene expression regulation.
www.jidonline.org S197